Microbial Inactivation for Safe and Rapid Diagnostics of Infectious Samples

Abstract

The high risk associated with biological threat agents dictates that any suspicious sample be handled under strict surety and safety controls and processed under high-level containment in specialized laboratories. This study attempted to find a rapid, reliable, and simple method for the complete inactivation of a wide range of pathogens, including spores, vegetative bacteria, and viruses, while preserving microbial nucleic acid fragments suitable for PCRs and proteinaceous epitopes for detection by immunoassays. Formaldehyde, hydrogen peroxide, and guanidium thiocyanate did not completely inactivate high titers of bacterial spores or viruses after 30 min at 21°C. Glutaraldehyde and sodium hypochlorite showed high microbicidal activity but obliterated the PCR or enzyme-linked immunosorbent assay (ELISA) detection of bacterial spores or viruses. High-level inactivation (more than 6 log 10 ) of bacterial spores (Bacillus atrophaeus), vegetative bacteria (Pseudomonas aeruginosa), an RNA virus (the alphavirus Pixuna virus), or a DNA virus (the orthopoxvirus vaccinia virus) was attained within 30 min at 21°C by treatment with either peracetic acid or cupric ascorbate with minimal hindrance of subsequent PCR tests and immunoassays. The data described here should provide the basis for quickly rendering field samples noninfectious for further analysis under lower-level containment and considerably lower cost.The high risk associated with biological threats dictates that any suspicious samples be handled under strict surety and safety controls and processed under high-level containment in specialized laboratories. These laboratories are complex, very expensive to operate, and need to be staffed by personnel from an extremely limited pool of experts. In addition, safe means of transporting samples suspected of containing highly virulent agents to specialized high-level containment laboratories for analysis also is expensive, requiring in many countries the custody of armed personnel. It can be estimated that several million dollars are spent annually worldwide to secure and safely transport an ever-increasing stream of suspicious biological samples, which are collected in theaters of war as well as in domestic environments.The objective of this study was to identify liquid chemical reagents with high efficiency to inactivate microbial organisms and viruses that, at the same time, would preserve the high sensitivity of subsequent diagnostics performed at a lower level of containment and cost.Considerable knowledge has been accumulated regarding the efficiency of microbicidal reagents and methods, particularly on inactivating bacterial spores in liquids, on surfaces, or in aerosols (4, 19), but the subsequent effect of these disinfectants on the performance of either nucleic acidbased or immunologically based diagnostics have received relatively little attention. We selected for study two aldehydes (formaldehyde and glutaraldehyde), a halogenating agent (hypochlorite), two peroxides (hydrogen peroxide and peracetic acid), an...

Photochemical Inactivation of Alpha‐ and Poxviruses

“…Review of previous photobiologic studies showed microbial inactivation results to be frequently impaired by lack of proper dosimetry or lacking controls that confounded results and prevented quantitative conclusions (reviewed in [25]). We took considerable effort in assuring that the viruses in this study were not inactivated by heat or direct germicidal UV radiation that could invalidate our photoianctivation experiments.…”

“…The quencher, a 6‐carboxy‐tetramethyl‐rhodamine (TAMRA), was located at the 3′end. The sequences of primers and probes for the amplification of the 145 bp vaccinia virus specific and the 192 bp pixuna virus‐specific amplicon, as well as the optimal cycling conditions, have been presented as supplemental information in a previous publication (25). The rtPCR targeted the A27L gene of poxviruses, the rtRT‐PCR targeted the nsP1 gene of Alphaviruses.…”

Laboratory Decontamination of HHS-Listed and HHS/USDA Overlap Select Agents and Toxins

“…Alkaline glutaraldehyde. Adjust the sample to pH 8 by adding sufficient sodium bicarbonate (Sagripanti et al, 2011) from a prepared stock. (For example, a 0.3% w/v final concentration of NaHCO 3 may be a useful starting point.)…”

“…Add household bleach to samples to a final bleach dilution of 1:10 to 1:100 v/v. Mix and incubate for 30 minutes (Sagripanti et al, 2011).…”

Biomodification Strategies for the Development of Antimicrobial Urinary Catheters: Overview and Advances

“…Ag alloy and Ag oxide have been used to create a layer in latex catheter which would lead to infection control during catherization . However, in a broader scene, Ag has been observed to prevent adhesion and growth of bacteria, E. coli and P. aeruginosa . Although, Ag salts may be directly used to coat the surface, nanosilver has recently gained enormous momentum toward the generation of an effective surface against microbes due to serious problem associated with metallic salts in terms of the leaching of the agent during use and hence leading to the toxicity of the tissue .…”